Drosophila functional screening of de novo variants in autism uncovers damaging variants and facilitates discovery of rare neurodevelopmental diseases

Marcogliese, Paul C.; Deal, Samantha L.; Andrews, Jonathan C.; Harnish, J. Michael; Bhavana, Venkata Hemanjani; Graves, Hillary K.; Jangam, Sharayu; Luo, Xi; Liu, Ning; Bei, Danqing; Chao, Yu-Hsin; Hull, Brooke; Lee, Pei-Tseng; Pan, Hongling; Bhadane, Pradnya; Huang, Mei‐Chu; Longley, Colleen M; Chao, Hsiao‐Tuan; Chung, Hyunglok; Haelterman, Nele A; Kanca, Oguz; Manivannan, Sathiyanarayanan; Rossetti, Linda; German, Ryan J.; Gerard, Amanda; Schwaibold, Eva Maria Christina; Fehr, Sarah; Guerrini, Renzo; Vetro, Annalisa; England, Eleina; Murali, Chaya N.; Barakat, Tahsin Stefan; Dooren, Marieke F. van; Wilke, Martina; Slegtenhorst, Marjon van; Lesca, Gaëtan; Sabatier, Isabelle; Chatron, Nicolas; Brownstein, Catherine A.; Madden, Jill A.; Agrawal, Pankaj B.; Keren, Boris; Courtin, Thomas; Perrin, Laurence; Brugger, Melanie; Roser, Timo; Leiz, Steffen; Mau‐Them, Frédéric Tran; Delanne, Julian; Sukarova-Angelovska, Elena; Trajkova, Slavica; Rosenhahn, Erik; Strehlow, Vincent; Platzer, Konrad; Keller, Roberto; Pavinato, Lisa; Brusco, Alfredo; Rosenfeld, Jill A.; Marom, Ronit; Wangler, Michael F.; Yamamoto, Shinya

doi:10.1016/j.celrep.2022.110517

Cited by 26 publications

(16 citation statements)

References 89 publications

(126 reference statements)

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“…This reward circuitry has been linked to distinct pathologies, ranging from autism spectrum disorder (ASD) to psychosis [81–85]. In agreement with a role for GlyRα2 to affect the circuitry involved in these pathologies were rare variants linked to ASD and neurodevelopmental disorders by large-scale next-generation sequencing studies [86–90]. Structure-function studies showed both loss, gain and altered function of GlyRα2 ASD mutations [89, 91].…”

Section: Discussionmentioning

confidence: 89%

Lack of the glycine receptor alpha 2 increases striatal activity and motivated behavior

Devoght

Comhair

Morelli

et al. 2022

Preprint

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Distinct developmental pathologies, including autism spectrum disorder and schizophrenia, exhibit impaired reward-motivated behavior. Key to proper reward-motivated behavior is the integration of dopaminergic and glutamatergic inputs to the striatum. The glycine alpha 2 receptor (GlyRα2) is the single functionally expressed glycine receptor in adult striatum, and is therefore ideally positioned to modulate striatal behavior and cellular signal integration. Here, we report excessive appetitive conditioning in GlyRα2 knockout animals. We next show that depletion of GlyRα2 enhances dopamine-induced increases in the activity of putative dopamine D1-expressing striatal projection neurons, while not affecting dopamine neuron activity. These in vivo behavioral changes are tied to enhanced striatal activation in GlyRα2 KO mice, since we found excessive locomotor responses to amphetamine in GlyRα2 KO mice that correlate with immediate early gene c-fos expression in the dorsal striatum. 3-D modeling revealed activation of cell ensembles in the striatum in response to D-amphetamine, an increase in number of cells but no shift in intercell distance histograms, in agreement with unaltered dopamine release, in GlyRα2 KO mice. Taken together, we show that depletion of GlyRα2 impairs reward-motivated behavior and altered striatal signal integration. This sheds important light onto the cellular mechanisms that underlie reward function, and pave the way towards novel therapeutics for the treatment of e.g. schizophrenia and autism spectrum disorder.

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Section: Discussionmentioning

confidence: 89%

Lack of the glycine receptor alpha 2 increases striatal activity and motivated behavior

Devoght

Comhair

Morelli

et al. 2022

Preprint

View full text Add to dashboard Cite

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“…Based on these results, we classify α2 T269M as a pathogenic alteration-of-function variant (given the reduced glycine EC 50 plus leak current) that is predicted to enhance glycinergic signaling in the developing brain. It is also noteworthy that the GlyR α2 T269M mutation has recently been reported as a de novo mutation in six additional female subjects (Marcogliese et al, 2022), making it the first recurrent GLRA2 pathogenic mutation. Using a novel Drosophila-based functional system for ASD mutations, Marcogliese et al (2022) also classified GlyR α2 T269M as a gain-of-function allele based on experiments overexpressing human GlyR α2 T269M in presynaptic photoreceptors and postsynaptic neurons, reporting a significant increase in amplitudes of "OFF" transients for the GlyR α2 T269M transgenic line.…”

Section: Discussionmentioning

confidence: 96%

Loss, Gain and Altered Function of GlyR α2 Subunit Mutations in Neurodevelopmental Disorders

Chen

Wilson

Schaefer³

et al. 2022

Front. Mol. Neurosci.

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Glycine receptors (GlyRs) containing the α2 subunit govern cell fate, neuronal migration and synaptogenesis in the developing cortex and spinal cord. Rare missense variants and microdeletions in the X-linked GlyR α2 subunit gene (GLRA2) have been associated with human autism spectrum disorder (ASD), where they typically cause a loss-of-function via protein truncation, reduced cell-surface trafficking and/or reduced glycine sensitivity (e.g., GLRA2Δex8-9 and extracellular domain variants p.N109S and p.R126Q). However, the GlyR α2 missense variant p.R323L in the intracellular M3-M4 domain results in a gain-of-function characterized by slower synaptic decay times, longer duration active periods and increases in channel conductance. This study reports the functional characterization of four missense variants in GLRA2 associated with ASD or developmental disorders (p.V-22L, p.N38K, p.K213E, p.T269M) using a combination of bioinformatics, molecular dynamics simulations, cellular models of GlyR trafficking and electrophysiology in artificial synapses. The GlyR α2V–22L variant resulted in altered predicted signal peptide cleavage and a reduction in cell-surface expression, suggestive of a partial loss-of-function. Similarly, GlyR α2N38K homomers showed reduced cell-surface expression, a reduced affinity for glycine and a reduced magnitude of IPSCs in artificial synapses. By contrast, GlyR α2K213E homomers showed a slight reduction in cell-surface expression, but IPSCs were larger, with faster rise/decay times, suggesting a gain-of-function. Lastly, GlyR α2T269M homomers exhibited a high glycine sensitivity accompanied by a substantial leak current, suggestive of an altered function that could dramatically enhance glycinergic signaling. These results may explain the heterogeneity of clinical phenotypes associated with GLRA2 mutations and reveal that missense variants can result in a loss, gain or alteration of GlyR α2 function. In turn, these GlyR α2 missense variants are likely to either negatively or positively deregulate cortical progenitor homeostasis and neuronal migration in the developing brain, leading to changes in cognition, learning, and memory.

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“…The number of epilepsy-related genes and variants will likely increase, and high-throughput screens are needed to meet the demand of functionally testing these variants. In a similar approach to the screen mentioned above, hundreds of variants regarding autism were functionally described in the fly ( Marcogliese et al, 2022 ). Patient variants were expressed under endogenous promoters of fly orthologues or overexpressed in different tissues.…”

Section: Drosophila Genetics and Tools To Investigate Seizur...mentioning

confidence: 99%

Drosophila melanogaster as a versatile model organism to study genetic epilepsies: An overview

Fischer

Karge²,

Weber³

et al. 2023

Front. Mol. Neurosci.

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Epilepsy is one of the most prevalent neurological disorders, affecting more than 45 million people worldwide. Recent advances in genetic techniques, such as next-generation sequencing, have driven genetic discovery and increased our understanding of the molecular and cellular mechanisms behind many epilepsy syndromes. These insights prompt the development of personalized therapies tailored to the genetic characteristics of an individual patient. However, the surging number of novel genetic variants renders the interpretation of pathogenetic consequences and of potential therapeutic implications ever more challenging. Model organisms can help explore these aspects in vivo. In the last decades, rodent models have significantly contributed to our understanding of genetic epilepsies but their establishment is laborious, expensive, and time-consuming. Additional model organisms to investigate disease variants on a large scale would be desirable. The fruit fly Drosophila melanogaster has been used as a model organism in epilepsy research since the discovery of “bang-sensitive” mutants more than half a century ago. These flies respond to mechanical stimulation, such as a brief vortex, with stereotypic seizures and paralysis. Furthermore, the identification of seizure-suppressor mutations allows to pinpoint novel therapeutic targets. Gene editing techniques, such as CRISPR/Cas9, are a convenient way to generate flies carrying disease-associated variants. These flies can be screened for phenotypic and behavioral abnormalities, shifting of seizure thresholds, and response to anti-seizure medications and other substances. Moreover, modification of neuronal activity and seizure induction can be achieved using optogenetic tools. In combination with calcium and fluorescent imaging, functional alterations caused by mutations in epilepsy genes can be traced. Here, we review Drosophila as a versatile model organism to study genetic epilepsies, especially as 81% of human epilepsy genes have an orthologous gene in Drosophila. Furthermore, we discuss newly established analysis techniques that might be used to further unravel the pathophysiological aspects of genetic epilepsies.

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Drosophila functional screening of de novo variants in autism uncovers damaging variants and facilitates discovery of rare neurodevelopmental diseases

Cited by 26 publications

References 89 publications

Lack of the glycine receptor alpha 2 increases striatal activity and motivated behavior

Lack of the glycine receptor alpha 2 increases striatal activity and motivated behavior

Loss, Gain and Altered Function of GlyR α2 Subunit Mutations in Neurodevelopmental Disorders

Drosophila melanogaster as a versatile model organism to study genetic epilepsies: An overview

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